Association of high LDL concentrations with erectile dysfunction from a Mendelian randomization study

Lipid metabolism plays a key role in erectile dysfunction. Our purpose was to evaluate the influence of lipid-lowering drugs on erectile dysfunction employing a two-sample Mendelian randomization (MR) study. Genetic instruments were employed to represent the exposure of lipid-lowering drugs. Inverse variance-weighted MR (IVWMR) was employed to calculate the estimation of effects. IVW-MR analysis showed that the positive relationship between the expression of HMGCR and the risk of erectile dysfunction (odds ratio [OR] = 1.27, 95% confidence interval [CI] 1.03–1.57; p = 0.028). No significant relationship was detected between NPC1L1, PSK9 expression and erectile dysfunction. This MR study suggested that HMGCR inhibitors are a more desirable treatment modality for patients with ED.


Genetic instrument selection
This investigation encompassed three classifications of FDA-endorsed lipid-lowering pharmaceuticals as exposure factors: HMGCR inhibitors, PCSK9 inhibitors, and the NPC1L1 inhibitor.
To corroborate the associations discerned through the employment of eQTLs as genetic instruments, we adopted a strategy that involved selecting SNPs situated within 100 kb windows surrounding each drug's target gene.These SNPs were associated with LDL cholesterol concentrations at a genome-wide significance threshold (p < 5.0 × 10 −8 ) and served as proxies for exposure to lipid-lowering medications.The GWAS summary data related to LDL cholesterol levels, procured from the Global Lipids Genetics Consortium (GLGC) and comprising a sample size of 173,082 participants, was employed to pinpoint these SNPs 19 (Supplementary Table S1).The screening flowsheet is shown in Fig. 1.
The selection of these genetic instruments was based on their association with LDL cholesterol concentrations at a genome-wide significance threshold (p < 5.0 × 10 -8 ).This criterion ensures that the selected SNPs are Table 1.Information of all GWAS summary data.robustly associated with the exposure.While each individual SNP might explain a small fraction of the variance in LDL concentration, the combined set of SNPs, especially considering their strong statistical association with LDL cholesterol (evidenced by their p-values), ensures a reliable genetic instrument.
The statistical evaluations comprised a primary analysis utilizing inverse-variance-weighted MR (IVW-MR), in addition to sensitivity analyses encompassing the F-Statistic, positive control assessment, linkage disequilibrium assessment, horizontal pleiotropy examination, heterogeneity testing, and MR-Egger regression.

Sources of outcome data
GWAS summary-level data concerning erectile dysfunction outcomes, sourced from the IEU GWAS database, which includes a sample size of 223,805, with 6175 cases and the remaining being controls 20 .

Statistical analysis
Primary MR analysis When utilizing genetic variants associated with LDL cholesterol concentrations as instruments, the IVW-MR method was implemented to amalgamate effect estimates.Allele harmonization and subsequent analysis were conducted using version 4.2.3 of the TwoSampleMR package within the R software environment.

Sensitivity analysis
We evaluated the strength of the SNPs used as instruments by calculating the F-statistic, including only SNPs with an F-statistic greater than 10 in order to mitigate the potential for weak instrument bias 21 .To validate both genetic instruments, we performed positive control analyses.Given that lipid-lowering drugs are known to effectively reduce LDL cholesterol levels, we investigated the relationship between the exposures of interest and LDL cholesterol levels as a positive control study for the eQTLs-based instruments.In the case of the instrument derived from the LDL cholesterol GWAS, we carried out a positive control study by examining the association between the exposures of interest and coronary heart disease.This was done because coronary heart disease is the primary indication for lipid-lowering medications.
In the IVW-MR method, we evaluated heterogeneity using the Cochran Q test, wherein a p-value below 0.05 indicates the presence of heterogeneity 22 .To assess the potential for horizontal pleiotropy among the SNPs utilized as instrumental variables, we utilized MR-Egger regression and Mendelian Randomization Pleiotropy RESidual Sum and Outlier (MR-PRESSO) analysis.In MR-Egger regression, the intercept term serves as a valuable indicator for assessing directional horizontal pleiotropy, whereby a p-value below 0.05 suggests the presence of horizontal pleiotropy 23 .MR-PRESSO analysis can identify horizontal pleiotropic outliers and offer adjusted estimates, with a p-value below 0.05 for the Global test indicating the presence of horizontal pleiotropic outliers 24 .All of the aforementioned analyses were conducted utilizing R software, version 4.2.3.

Analysis using the two-sample MR
IVW-MR analysis results showed that the positive relationship between the expression of HMGCR and the risk of erectile dysfunction (odds ratio [OR] = 1.27, 95% confidence interval [CI] 1.03-1.57;p = 0.028), suggesting that lower HMGCR expression reduced the risk of erectile dysfunction (Fig. 2 and Table 2).No significant relationship was detected between NPC1L1, PSK9 expression and erectile dysfunction.

Sensitivity analysis
A lack of causal association remained in all sensitivity analyses (all p > 0.05).Likewise, there was no clear evidence of heterogeneity (all p > 0.05) or pleiotropy (all p values for intercept > 0.05) regarding HMGCR, NPC1L1 and PCSK9 in erectile dysfunction and positive control coronary heart disease (Table 3).Moreover, the leave-oneout analysis plot results certified that the above results were unchanged by the removing of any SNP and were quite robust (Fig. 3).

Discussion
In the present study, increased HMGCR gene expression was associated with an increased risk of erectile dysfunction, suggesting that HMGCR inhibitors may reduce the risk of erectile dysfunction.The expression of NPC1L1 and PCSK9 was not significantly associated with ED.
Statins are the most commonly used HMGCR inhibitors in clinical practice and play an important role in lowering LDL and cholesterol and slowing the progression of atherosclerosis 25 .Increased production of LDL receptors is thought to be the main mechanism driving the action of statins 26 .Statins also have an ameliorating effect on platelet and vascular endothelial function 27 .However, statins still have some side effects, including statin-related muscle symptoms, new-onset type 2 diabetes, neurological and neurocognitive effects 28 .
Statins may improve penile erectile function in several ways; a reduction in LDL may improve endothelial function, and good endothelial function is important for penile erection.Moreover, statins may increase the availability of nitric oxide, which plays a key regulatory role in penile erection and resistance to oxidative stress.However, statins may also impair penile erectile function because they block reductase in the early stages of cholesterol biosynthesis, thereby reducing testosterone formation 29 .Testosterone is closely associated with sexual behavior, and it enhances the overall male sexual response 30 .In patients with reduced testosterone levels, their sexual desire and number of morning erections are significantly reduced 31 .
However, the effect of statins on ED in men remains controversial in current clinical studies.In 1812 men treated with statins, no association between statin use and the development of ED was found by testing testosterone and luteinizing hormone 32 .In contrast, in a study by BRUCKERT et al. patients who were treated with   statins were more likely to suffer from ED 33 .And in another individual study, it was also found that some patients experienced decreased libido and significantly lower testosterone after taking different types of statins 34 .These opposite findings may be due to various differences in the age of initiation of statin therapy, the dose used, and the type of their underlying disease.
As for NPC1L1 and PCSK9 inhibitors, there was no significant correlation between them and ED in this study.We consider that on the one hand, it is because the lipid-lowering effect of NPC1L1 inhibitors is not as strong as that of statins; and PCSK9 inhibitors, although they have a strong lipid-lowering effect, have a low bioavailability and are expensive, which leads to the fact that both of them are usually used only by patients with poor lipids 35,36 .This problem of poor lipids may make the protective effect of NPC1L1 and PCSK9 inhibitors on ED www.nature.com/scientificreports/not significant.On the other hand, the inhibition of the cholesterol biosynthesis process by statins may inhibit multiple actions of downstream products together, which may also be a factor in the ability of statins to exert ED protection, but this needs to be verified by more subsequent studies.

Strengths and limitations
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Conclusion
In the present study, we reduced the risk of confounding by a Mendelian randomization study and found that HMGCR gene expression was positively associated with ED occurrence, whereas NPC1L1 and PCSK9 gene expression did not correlate with ED occurrence.It is suggested that the use of HMGCR inhibitors may reduce the occurrence of ED and that HMGCR inhibitors are a more desirable treatment modality for patients with cardiovascular disease combined with ED.

value for MR-Egger intercept MR Egger P-value for Cochran Q test Inverse variance weighted P-value for Cochran Q test P value for MR-PRESSO global test
1) Participants of European ancestry only were enrolled in the study; future studies of populations of other ethnicities are needed to test the generalizability of the current study's findings.(2)Weutilized MR to reduce confounding in studying ED's association with lipid-lowering drugs.While stringent SNP selection and sensitivity analyses bolster our study, it's essential to acknowledge that potential biases, like horizontal pleiotropy, can't be fully eliminated.(3) Concerns arose over potential overlaps between IEU GWAS and our eQTL-derived GWASs.Despite verifying distinct cohorts, minor overlaps are possible.This might introduce biases, but our MR-Egger and MR-PRESSO analyses aim to control such biases.(4) Our MR analysis carefully chooses SNPs based on proximity to drug target genes and LDL cholesterol significance.While methods like the Cochran Q test help, the intrinsic variability remains.Despite robust techniques like IVW-MR and MR-Egger regression, potential biases from SNP variability persist.While our results offer valuable insights, they warrant cautious interpretation, suggesting future research could benefit from expanded SNP criteria for enhanced robustness.